Borrelia burgdorferi (Bb), the causative agent of Lyme disease, is the most common arthropod-borne infectious agent in the United States and contributes to significant morbidity resulting in extensive health care costs. Bb is transmitted to humans and other mammalian hosts following the bite of infected Ixodes spp. ticks. A number of studies have shown the importance of rpoS as a central regulator of gene expression in Bb that facilitates the adaptation of the spirochetes to vertebrate-host specific conditions. An unanswered question in the field of borreliosis is how rpoS is repressed to facilitate adaptation of Bb to vector-specific conditions. An open reading frame in B. burgdorferi, BB0693, annotated as XylR1 (Xylose utilization Regulator 1), was identified as a DNA - binding protein and bound to up-stream regions of rpoS. Deletion of this regulator in a virulent strain of Bb resulted in de-repression of rpoS and its regulon. Based on additional in vitro and in vivo characterization, BB0693 was designated as Borrelia host-adaptation Regulator (BadR). The central hypothesis is that BadR regulates rpoS to facilitate adaptation of Bb to different hosts. Critical genetic and biochemical tools such as badR deletion mutants and cis-complemented strains restoring a functional copy of badR carrying all the infection-associated plasmids have been generated. The central hypothesis will be tested with two Specific Aims. In specific aim 1, the regulatory mechanisms of BadR will be determined. The working hypothesis is that BadR regulates levels of rpoS and other open reading frames in response to host-specific signals/nutrients. In this aim the following will be determined: 1) the effects of PTS/non-PTS sugars on growth of badR mutants; 2) the regulatory effects of sugars on transcriptional and protein profiles of Bb and 3) the membrane localization of BadR in response to different sugars. Completion of this aim will provide the molecular basis of how BadR regulates gene expression in response to different sugars and its effects on patho-physiology of Bb. In Specific Aim 2 the role of BadR in the vector and mammalian phases of Bb infection will be determined. The working hypothesis is that BadR is essential for the vector and mammalian phases of Bb infection. In this aim the following will be determined: 1) the transcriptional levels of badR and badR-regulated genes in infected Ixodes scapularis ticks; 2) the regulatory effects of badR in artificially fed- ticks and 3) transcriptional levels of badR and its regulon in infected C3H/HeN mice. These studies will help determine the role of BadR in the host-specific adaptation of Bb and for the completion of tick-mouse-tick cycle of B. burgdorferi.